University Of Minnesota

NIH Research Projects · FY 2026 · 2023-06

Summary Alzheimer's Disease and Related Dementias (ADRD) is an overwhelming medical condition at any age, but ADRD in younger adulthood is particularly devastating, affecting quality of life and independence of individuals in their prime years. Early onset ADRD (EOD) is defined as an ADRD diagnosis before age 65. While it is commonly perceived that EOD occurs primarily as a rare genetic syndrome, the known genetic variants account for less than 5% of cases. Despite the distressing course and unclear nature of the disease in the majority of cases, EOD is widely understudied. Current data on the prevalence and incidence of EOD seem to underestimate the magnitude of the problem and there is no information available regarding predisposing and/or protective factors for EOD. Using the infrastructure of an international Dementia Risk Pooling Project (DRPP), we propose a prospective study of EOD development which comprises individuals from five large multi-ethnic population-based cohort studies (ARIC, MESA, FHS, Whitehall II and, UK Biobank). This study provides the opportunity to (1) refine estimates of incident EOD, (2) investigate the role of cardiovascular, lifestyle and behavioral risk factors in the onset of EOD and (3) study whether a favorable midlife risk profile in the presence of genetic predisposition delays EOD age of onset. This evaluation will be the first study to pool multiple international population-based cohorts to prospectively study ADRD before the age of 65 in young and middle-aged adults. The current notion that EOD is merely driven by genetic syndromes has shadowed efforts to identify distinct predisposing and/or protective factors for EOD and targeting vulnerable populations for early detection and prevention. The findings will shed much needed light on the vulnerability and unique risk factors for EOD and may lead to development of more effective targets for prevention to delay onset and progression of disease. EOD is relatively rare and thus primordial and primary prevention may be more efficient than screening and secondary or tertiary prevention. Our data on EOD risk factors will strengthen targeted intervention strategies with focus on primordial and primary prevention.

NIH Research Projects · FY 2024 · 2023-06

ABSTRACT BRAIN Initiative-funded, large-scale approaches to classify neurons based on transcriptomic, morphological and electrical properties have unveiled dozens of unique cell classes in the mouse brain. However, whether they represent functionally diverse populations of relevance to animal perception and behavior remains an open question. Dissecting their individual roles requires the integration of targeted recording techniques and optogenetic manipulation approaches, which operate at the physiologically relevant spatiotemporal scales (i.e. cellular resolution and millisecond timescales). Here, we propose to use high-speed (0.4-1 kHz), genetically encoded fluorescence voltage imaging to understand the role of distinct interneuronal populations in attentional modulation of visual processing during visually guided behavior. First, we will establish the optical instrumentation for high-speed, dual channel voltage imaging with non-overlapping structured illumination. We will further validate the use of our second-generation, fluorescence resonance energy transfer (FRET)-opsin indicators Ace-mNeon2 and VARNAM2 and their reverse response polarity variants pAce and pAceR, for concurrent voltage recordings from pairs of interneuronal ensembles and pyramidal neurons in awake, running mice. Thereafter, using simultaneous triple-population voltage imaging, we will assess the effects of attention on the firing rates of the three cell classes during visually guided behavior and compute the spatiotemporal correlations in the activation patterns of neighboring neurons. Separately, we will measure the visual tuning properties of the same neurons during presentations of drifting grating stimuli. We will further draw a correlation between neuronal attention modulation index and feature selectivity to test the applicability of the feature similarity gain model. Lastly, to establish causal roles in the attentional modulation of visual responses, we will optogenetically manipulate the activity of select interneurons in a spatially precise manner, while recording the voltage responses in neighboring pyramidal cells when mice are engaged in the behavioral task. Our proposed work will (1) elucidate the role of distinct interneuron-types in visual attention and enable functional cross-comparisons within the same animals and at increased spatiotemporal resolution; (2) uncover synergistic and antagonistic relationships between neighboring pyramidal neuron-interneuron pairs for neurons that are positively versus negatively modulated by attention; (3) test the applicability of the feature similarity gain model in rodents and (4) establish causal roles for distinct interneuronal populations in attentional modulation of visual processing. Together, our work will establish simultaneous, multipopulation voltage imaging as the preferred modality to unravel the real-time functional differences between neuron-types in perception and behavior.

NIH Research Projects · FY 2025 · 2023-06

Abstract: Epstein-Barr virus (EBV) associated diseases remain a huge burden in human health. As an orally transmitted pathogen, EBV infection causes infectious mononucleosis and ~200,000 cases of various cancers, including nasopharyngeal carcinoma that occurs in a space immediately adjacent to the oral cavity, some B cell malignancies, and ~10% of gastric cancer. In HIV infected people, EBV causes oral hairy leukoplakia of tongue. To understand the molecular mechanisms through which EBV contributes to disease development, EBV-transformed lymphoblastoid cell lines are used as a model system. EBV nuclear antigen leader protein (EBNALP) is essential for EBV to transform naïve B lymphocytes. Most of its known functions are linked to EBV transcription activator EBNA2. However, EBNALP binds to many enhancer and promoter sites independent of EBNA2. Perturbations of these EBNALP sites with CRISPRi significantly decreased these enhancers’ linked gene expression. Little is known about how EBNALP exerts its EBNA2 independent functions. It is also not known how EBNALP is tethered to the enhancer/promoter sites and how they affect transcription. Therefore, we hypothesize that EBNALP exploits host transcription programs to gain access to host enhancers/promoters, and contributes to EBV transformation through EBNA2-independent mechanisms. During my mentored period, I will address the fundamental question of how EBNALP binds to DNA. We will use CRISPR-based assays to identify host proteins essential for EBNALP enhancer activation. I will first focus on sequence-specific transcription factors (TFs). Chromatin immune precipitation (ChIP) based assays will be used to test the effects of knockout on EBNALP DNA binding. During my R00 phase, I will perform research independently and distinguish my work from my mentor’s by studying different aspects of EBNALP. I will focus my studies on characterizing the enhancer protein complexes assembled by EBNALP onto the enhancers to regulate transcription activity. I will focus on transcription cofactors, basal transcription factors, and histone modifying enzymes. Understanding the mechanisms through which EBNALP binds to DNA and regulates gene transcription may provide promising targets for treating EBV-associated diseases.

NIH Research Projects · FY 2026 · 2023-06

PROJECT SUMMARY Cellular energy metabolism is a fundamental process of life that produces biochemical energy in the form of adenosine triphosphate (ATP) to support neuronal activity and brain function. Glucose and oxygen are the main energy substrates of the brain and are metabolized through glycolysis, the tricarboxylic acid cycle and oxidative phosphorylation pathways, constituting a neuroenergetic network that effectively regulates ATP production and homeostasis. ATP production and homeostasis are affected when brain states change, as signs of altered cerebral glucose and oxidative metabolism are commonly seen in aging, neurodegenerative diseases, psychiatric disorders, stroke and cancer. Despite the important roles of brain energy metabolism, metabolic alteration and reprogramming in health and disease, noninvasive neuroimaging tools capable of mapping and quantifying key features of neuroenergetic network in the human brain are still lacking. Over the past two decades, we have developed three ultrahigh-field (UHF) metabolic imaging techniques based on deuterium-2 (2H), oxygen-17 (17O), and phosphorus-31 (31P) magnetic resonance spectroscopy (MRSI) imaging capable of noninvasive and quantitative assessment of brain energy metabolism along major metabolic pathways. However, X-nuclear MRSI-based methods face severe challenges in translational applications due to low detection sensitivity and metabolite content, and prolonged scanning time. This project aims to develop and integrate multiple cutting-edge technologies to build next generation high- resolution, high-performance and translatable neuroimaging tools on an FDA-approved 7 Tesla clinical scanner for quantitatively imaging key metabolic rates and other essential neurophysiological parameters related to energy metabolism in healthy and diseased human brains. Three pilot studies are proposed to test and demonstrate the utility and feasibility of the novel neuro-metabolic imaging tools to quantitatively study neuroenergetics and metabolic reprogramming in brain activation, aging processes and brain tumors, aiming to understand their critical roles in brain function and disease. This project leverages the interdisciplinary expertise of an outstanding team leading in the research field, excellent imaging facilities and resources, and close collaboration among team members. The advanced neuroimaging tools established by this project is expected to have significant impact on changing the paradigm of neurometabolic imaging and energy metabolism research, and enable translational studies of human brain bioenergetics and metabolic reprogramming under physiopathological conditions.

NIH Research Projects · FY 2024 · 2023-06

Abstract Bone marrow perfusion can provide essential knowledge about bone physiology, improve our understanding of disease etiology and pathophysiology, assist the differentiation between normal and abnormal bone marrow, and assess the response to prescribed therapies. Arterial spin labeling (ASL) magnetic resonance imaging (MRI), as a noninvasive and non-contrast-enhanced approach, is well suited for longitudinal monitoring of disease progression and routine evaluation of therapy response. But as revealed by our recent studies, there exist technical challenges (e.g., low signal-to-noise (SNR) ratio due to significantly low perfusion in epiphyseal yellow bone marrow mainly consisting of fat cells with a sparse network of capillaries), and methodological limitations (e.g., single slice coverage and impractically long acquisition time for multi-slice imaging at 3T), hampering routine application of ASL imaging in the knee. Ultrahigh (≥7T) magnetic field can specifically benefit ASL imaging and overcome these challenges by increasing SNR, prolonging blood and tissue T1, and improving parallel imaging performance. However, the current clinically approved single-transmit MRI system and associated imaging methods are incapable of managing the transmit B1 (B1+) fields needed to realize the promised improvement in imaging quality, reliability, and robustness of 7T while existing RF coils are unable to provide adequate B1+ coverage for optimal ASL imaging. Our long-term goal is to develop and improve UHF imaging methods to better facilitate scientific research and clinical studies to improve the management of skeletal diseases and patient healthcare. The rationale is that the existing technical challenges for UHF imaging, including ASL, can be overcome or mitigated, ultimately to realize UHF potentials with promised benefits and superior imaging ability. The objective of this proposal is to develop an optimized, safe, and efficient pTx platform for knee ASL imaging (Aim 1), to develop and optimize pTx-integrated knee ASL imaging methods (Aim 2) and to explore their clinical potentials via studies with a cohort of JOCD patients, an ideal testbed for the developed methodologies. Accomplishing these aims will enable us to fully explore and utilize pTx potentials to overcome the existing technical challenges for knee ASL imaging at 7T. Once developed and optimized, the ASL methods, along with the pTx knee imaging platform, will provide clinicians and research community a powerful tool to enrich our insights into disease etiology and pathophysiology and improve the management of not only JOCD but also various impactful common knee diseases (e.g., osteoarthritis). Novel knowledge and experience from our pioneering development will speed up the maturity of new technology toward early approval of the next generation of clinical UHF MRI systems.

NIH Research Projects · FY 2025 · 2023-06

Existing neurotechnologies continue to make significant clinical impact, but challenges remain for scientists and engineers in moving new devices from the bench to the bedside. Over the past two years (through an R25 grant), we developed a comprehensive, freely-available online short course on neurotechnology translation and commercialization called the NeuroTech Course featuring 20 self-paced video lectures from a diverse and experienced group of program faculty, topical questions, and a growing catalog of online resources for more advanced learning. These short-course video lectures cover preclinical model systems, safety and efficacy studies, good laboratory practices, device testing, quality system processes, regulatory agency interactions, steps in developing an investigational device exemption (IDE) application, reimbursement agency interactions, clinical trial design with an emphasis on quantitative outcome measures of target engagement, bioethical considerations that are specific to neural medical devices, techniques for securing strong intellectual property claims, funding opportunities available for technology development and clinical trials, and advice on moving neurotechnology into successful commercial ventures. With this grant, we propose to significantly amplify this course by (1) creating virtual “sprints” in which program faculty guide participant cohorts through the online lectures (to maximize adherence to the curriculum and enable participants to ask follow-up questions along the way). These online “sprints” will then be coupled with a (2) 3-day hands-on workshop (in person and virtual options) in which participants will work with program faculty to review case studies; discuss best practices through a series of intensive exercises taking a clinical need through the innovation process; and be mentored through the development of their own plans. The course will target senior postdoc scholars; academic and clinical faculty; and start-up company scientists, engineers, and entrepreneurs who are preparing to develop research grant proposals with a clinical component. This course will serve as an important bridge for early-stage entrepreneurs to be successful in moving to later-stage programs, including NIH C3i, NSF I-Corps, incubators, and accelerators.

NIH Research Projects · FY 2026 · 2023-06

Project Summary The principal investigator Dr. Lakshmi’s goal is to develop evidence-based technology interventions that support chronic disease care in older persons including those with mild cognitive impairment (MCI) and early stage Alzheimer’s disease and Alzheimer’s Disease Related Dementia (AD/ADRD). This K24 application will provide her with protected time to: 1) build skills in implementation science, aging research and the conduct of research with persons with cognitive impairment and their family caregivers; 2) accelerate mentoring of early career clinical faculty in the Department of Neurology particularly on K-awards and transition to the 1st R01; 3) obtain pilot feasibility and effect size data for a larger clinical trial of a mobile health technology (mHealth) intervention to improve hypertension (HTN) care in persons with MCI and early stage AD/ADRD. Dr. Lakshmi is a tenured faculty in the Division of Epidemiology and Community Health, School of Public Health (SPH) at the University of Minnesota and in the Department of Neurology. Her rich institutional environment includes partnerships with the Divisions of Biostatistics and Health Policy and Management, Medical School departments and community health systems. Dr. Lakshmi is a member of the Center for Health Aging and Innovation, a collaboration that aims to advance interdisciplinary aging research. The research focus of this K24 is improving the management of uncontrolled HTN in persons with MCI and early stage AD/ADRD. HTN is the most significant stroke, cardiovascular disease and dementia risk factor and is substantially under-treated especially in older persons. Dr. Lakshmi has worked on mHealth interventions since 2011, with a successful record of multiple funded grants. Her clinical trial, mGlide RCT, is implementing a mHealth-based care model for HTN care in community health systems serving diverse, low-income patients. In this K24, Dr. Lakshmi will develop mGlide-Care – an adaptation of mGlide – to address uncontrolled HTN in people with MCI and early AD/ADRD. Aim 1 will engage stakeholders to study the acceptability of mHealth mediated HTN care and will use their input to develop mGlide-Care. Stakeholders are persons with early stage AD/ADRD and MCI, unpaid family caregivers, primary care providers, geriatricians and clinical pharmacists. Aim 2 is a feasibility pilot to test mGlide-Care vs. usual care in 75 participants with uncontrolled HTN and early stage AD/ADRD or MCI. Caregivers will assist participants. Outcomes will include HTN control and participant and caregiver reported measures. Dr. Lakshmi is the site-PI for two NIH research networks, StrokeNet and DISCOVERY (Determinants of Incident Stroke Cognitive Outcomes and Vascular Effects on RecoverY). She also has access to rich cohort data including Atherosclerosis Risk in Communities-Neurocognitive study (ARIC-NCS) and others. Dr. Lakshmi will leverage the substantial resources from her health technology grants, StrokeNet, DISCOVERY, and cohorts including ARIC-NCS to mentor junior investigators in NIA priority areas. Her goal is to prepare her mentees for their own major scientific advances and independent research careers.

NIH Research Projects · FY 2026 · 2023-05

PROJECT SUMMARY/ABSTRACT Prenatal alcohol use (PAU) is associated with increased likelihood of obstetric complications, including preterm labor, miscarriage, and stillbirth, and is also the direct cause of Fetal Alcohol Spectrum Disorders, a collection of neurodevelopmental disorders that cause lifelong neurobehavioral and craniofacial abnormalities. In 2020, the Center for Disease Control and Prevention reported that approximately 1 in 7 pregnant women had consumed alcohol in the past month, which is in line with increasing trends since 2011. Thus, PAU is a major and growing public health problem, so it is imperative to understand the risk factors for PAU to inform prevention and intervention. Stress is one such key risk factor. Pregnant women with frequent stress have a 3- fold higher risk of binge drinking than pregnant women without frequent stress. Furthermore, pre-pregnancy alcohol use is consistently associated with PAU, suggesting that understanding the etiology of alcohol misuse outside of pregnancy is essential for preventing PAU. Correspondingly, stress plays a crucial role in Alcohol Use Disorder (AUD) in women outside of pregnancy, as women are more vulnerable to relapse following stressful triggers relative to men. This vulnerability may in part be driven by sex/gender (SG) differences in neurocircuitry related to processing stress in AUD. Increased stress vulnerability has been linked to dysfunction in the “salience network” (SN), which is a collection of brain regions, primarily the insula, the dorsal anterior cingulate cortex, and inferior parietal lobule, that responds to salient, potentially stressful stimuli and is also highly reactive to substance use cues, including alcohol. Furthermore, social support may serve as a resilience factor against stress-related alcohol misuse, particularly in women. If and how social support can also buffer the brain’s heightened vulnerability to stress in AUD, and if there are SG differences in this effect, remains to be examined. Moreover, whether stress and social support similarly affect PAU is unclear. My overarching hypothesis is that women, relative to men, are more vulnerable to stress-related alcohol misuse via enhanced SN reactivity; however, social support will have a stronger buffering effect on this relationship in women. The specific aims of this project are to (1A) assess SG differences in the role of the SN on the relationship between stress reactivity and alcohol use levels in people with AUD, (1B) assess SG differences in whether social support protects against alcohol misuse by altering stress-related SN reactivity, and (2) determine risk and protective factors for prenatal alcohol use. Achieving these goals will inform scientifically- grounded treatments for AUD in women during and outside of pregnancy, as well as prepare me for a successful career as a physician-scientist in academic medicine.

NIH Research Projects · FY 2026 · 2023-05

Cardiovascular diseases are both common and deadly. For example, peripheral artery disease affects more than 10M Americans resulting in more than 150,000 limb amputations each year in the U.S. In addition, more than 300,000 patients have coronary artery bypass grafting (surgical revascularization). Current medical therapies for vascular disease include limb amputation and vascular bypass grafting--these therapeutic interventions have significant limitations. These diseases are chronic, debilitating, lethal and they warrant new and novel therapies. Previous studies have demonstrated the essential role for pioneer factors that modulate chromatin accessibility and thereby impact the binding of early transcriptional regulators for lineage specification. We have recently demonstrated that ETV2 is an essential pioneer factor for endothelial, vascular and blood lineages. We have used global and conditional gene disruption strategies, fate-mapping, gene editing, single cell RNA-seq, ATAC-seq and ChIP-seq assays to provide supportive data for this application. In addition, we defined an important ETV2-miR130a-PDGFRa cascade that governs endothelial development. Furthermore, our recent publications and our preliminary data support the overall hypothesis that ETV2 is a pioneer factor that regulates the specification of the endothelial lineage. In these proposed studies, we will use a number of unique genetic models that we have engineered and we take an innovative strategy to define the mechanisms whereby ETV2 functions as a pioneer factor to regulate cardiovascular regeneration. To examine our hypotheses, we will address the following specific aims: Specific Aim #1: Specific Aim #1: To further define the mechanisms whereby ETV2 functions as a pioneer factor during embryogenesis and reprogramming to the endothelial lineage; Specific Aim #2: To define the role of chromatin modifying factors and ETV2 during embryogenesis and reprogramming to the endothelial lineage and Specific Aim #3: To examine the factors that promote ETV2 mediated reprogramming of the endothelial lineage in vitro and in vivo. These aims will utilize our recently engineered genetic mouse models, ATAC-seq, MNase-seq, ChIP-seq, inducible mouse model, cardiac injury model in the adult mouse, novel and bioinformatics algorithms to comprehensively define the mechanisms whereby ETV2 functions as a pioneer factor and will serve as prelude for therapeutic initiatives to engineer and promote regeneration of the cardiovascular lineages. Given the tremendous morbidity and mortality of cardiovascular disease in our society, the potential impact of this proposal is significant.

NIH Research Projects · FY 2026 · 2023-05

PROJECT SUMMARY The pathogenesis of Alzheimer's disease (AD) remains elusive. Inheritance of the apolipoprotein (APO) E4 allele is the strongest genetic risk factor for sporadic late onset AD, whereas the APOE2 allele is protective and the most common APOE3 allele is neutral. While the mechanisms by which APOE4 modifies the risk of AD are not fully elucidated, compelling evidence indicates that the pathogenic effects of APOE4 are mediated by lipid- related pathways. Integrative multi-omics studies have consistently demonstrated the strong association of lipid pathways with AD phenotypes and that APOE4 disrupts intra/intercellular lipid homeostasis in cellular, organoid, and animal models as well as postmortem brain tissue from individuals carrying different APOE alleles with or without AD. Intriguingly, emerging evidence suggests that specific species of lipids/metabolic alterations in subcellular organelles, in particular mitochondria, lead to neurotoxicity and neurodegeneration. Mitochondria are the powerhouse of cells and provide the energy to sustain vital cellular functions. Notably, brain contains two major populations of mitochondria, the synaptic mitochondria that originate from the synaptic bouton of neurons and the non-synaptic mitochondria that originate from neuronal and glial cell bodies. Lipidomic analysis indicates that synaptic and non-synaptic mitochondria have distinct lipid profiles that regulate compartmental energy metabolism in the brain. Importantly, dysfunction of mitochondria, especially synaptic mitochondria, is well established as one of the earliest deficits in the progression of AD. APOE4 has been associated with increased impairment of mitochondrial structure and function compared with APOE3 in various models and human patients. However, whether APOE genotypes regulate the lipidome of mitochondria during brain aging and whether the dynamic changes of mitochondrial lipidomes affect the progression of AD are unknown. We hypothesize that mitochondrial lipidomic dynamics and its interaction with APOE4 drive pathogenic brain aging and AD. Three independent yet interrelated specific aims are proposed to test the hypothesis, using both mouse models and human brains and a combination of behavioral and pathological approaches, coupled with innovative targeted and unbiased cellular, molecular technologies, including lipidomics, transcriptomics, and brain clearing and 3D imaging. Aim 1 is to assess the impact of APOE isoforms on mitochondrial lipidomic dynamics associated with brain aging in humanized APOE4 and APOE3 mice. Aim 2 is to elucidate the relationship between mitochondrial lipidomic dynamics and the progression of cognitive deficits and amyloid pathology in APP/PS1 mice. Aim 3 is to define the interaction of mitochondrial lipidome with different APOE isoforms and its relation to cognitive function and AD pathology in human brains. The results are expected to uncover the impact of mitochondrial lipidomic dynamics and its interaction with APOE on brain aging and AD, and identify novel targets/pathways that can be harnessed as diagnostic biomarkers and/or for therapeutic development to defeat AD.

NIH Research Projects · FY 2026 · 2023-05

PROJECT SUMMARY/ABSTRACT Tourette Syndrome (TS) is a chronic, childhood-onset neurodevelopmental disorder that affects 1-3% of people and is associated with adverse functional impacts. TS is characterized by tics, which are involuntary, repetitive movements and vocalizations. A current challenge in clinical care for TS is the lack of objective, quantitative, scalable tools to measure tics for the purposes of diagnosis and symptom severity monitoring. The overall objective of the proposed study is to use video-based methods in a large, diverse community sample to inform quantitative and automated phenotyping of tics. This study builds on prior work, including: 1) video-based observational methods with trained human raters to quantify tics for research purposes, 2) computer vision and machine learning techniques for movement analysis and medical diagnostic aids, and 3) preliminary data indicating supervised learning methods can be used to automate detection of eye tics with high accuracy. In Aim 1, videos and clinical data from N = 1,000 individuals with tics will be collected using remote and internet-based methods. A deep phenotyping approach will be used to quantitatively describe the phenotypic spectrum of observable motor and vocal tics, empirically derive tic severity benchmarks, and identify patient subgroups. In Aim 2, our computer vision team will apply supervised machine learning methods to Aim 1 data to create an algorithm capable of detecting the most common tics. Aim 3 will prospectively test the Aim 2 algorithm in N = 60 patients who completed TS treatment in a separate clinical trial to establish the algorithm’s sensitivity to change and convergent validity with current gold-standard tic severity measurement. This project will enable us, for the first time, to quantify the spectrum of observable tics in a large community sample, knowledge that will have immediate clinical relevance for diagnostic decision making and patient education. Aim 2 will yield a computer algorithm capable of autonomously quantifying the most common motor tics, a critical next step toward developing accurate, clinically valid, and scalable assessments for tic screening, diagnosis, treatment decision making, and symptom quantification in clinical trials.

NIH Research Projects · FY 2026 · 2023-05

This proposed study concerns the proteolytic machinery that regulates tissue aging, inflammation, and metabolic deregulation. The proteasome population is largely shifted from the standard proteasome to the immunoproteasome under inflammation. The immunoproteasome is different from the standard proteasome in the 20S core three catalytic subunits. It has distinct processing kinetics and cleavage preferences, and associates with the 11S regulatory complex to perform ATP- and ubiquitination-independent degradation of proteins. Its expression is highly increased in high fat diet mouse liver and in liver biopsies from patients who have chronic active hepatitis or cirrhosis. The immunoproteasome level is highly increased in pathological hepatocytes from human in direct correlation with the histopathological grade of inflammation. The immunoproteasome is highly induced in aged mouse and rat tissues. Its prolonged presence can cause chronic inflammation and tissue damages and can have devastating effects on the stability of proteins, many of which are anti-inflammatory factors or homeostatic factors otherwise stable. Our study is based on the overarching concept that the immunoproteasome plays the major role in inflammation-specific degradation of proteins. The degradation may cause metabolic reprogramming and the transition between glycolysis and oxidative phosphorylation, shifting the cellular metabolism that fits in the inflammatory conditions. The long- term goal of our study is to understand the functions of the immunoproteasome in inflammatory metabolic diseases. Its strong relevance with inflammation, its immunological aspect of regulation, its dynamic nature, and its high expression in immune cells and aged tissues and high fat diet mouse tissues all point toward the possibility that the immunoproteasome might play crucial roles in inflammatory metabolic diseases. The objective of this proposed study is to define the mechanisms by which the immunoproteasome regulates energy metabolism via the selective digestion of key metabolic regulators. Despite its discovery nearly three decades ago, its specific function remains largely unknown other than its role in producing antigenic peptides. We have identified several regulators of the mitochondrial biogenesis and metabolism under the control by the immunoproteasome. We hypothesize that the immunoproteasome induces metabolic reprogramming by digestion of proteins involved in metabolic homeostasis. Although we do not exclude the possibility that the reprogramming might be protective in early stages of stress response, we hypothesize that its excessive, prolonged presence disturbs metabolic homeostasis. This grant will focus on the 11S-associated immunoproteasome, the major type of the proteasome during inflammation. We will pursue three specific aims. First, we will determine how the immunoproteasome perturbs mitochondrial metabolism using mouse and cellular models. Second, we will define the role of PA28alpha-associated immunoproteasome in hepatic metabolism. Third, we will identify the molecular factors that mediate 11S-immunoproteasome functions.

NIH Research Projects · FY 2026 · 2023-05

Project Summary Candidate and Career Goals: I intend to become an independent scientist at a research-first academic institution, bridging across levels of description (i.e., from computations to neurons) and furthering our understanding of how the brain infers the world around us, as well as ourselves within it. I trained as a cognitive scientist (winning the Glushko Prize for best dissertation in Cognitive Science, worldwide), with a focus on understanding our sense of self-location; where am “I” located in space. I am now training in systems neuroscience, developing expertise in large-scale rodent neurophysiology and with a focus on the dynamic aspects of self-location; spatial navigation. These experiences complement each other; from behavioral computations to single-units, and from static to dynamic self-location. Environment and Career Development Plan: I am mentored by Dr. Dora Angelaki (NYU, expertise in navigation) and co-mentored by Drs. David Schneider (NYU, rodent self-generated actions) and Cristina Savin (NYU, data science). Further, I am a scientist member of the International Brain Lab, allowing me the opportunity to leverage world-class expertise (22 labs) in rodent neurophysiology. My training during the K99 will focus on model-based analyses of behavior and neurons during continuous and complex naturalistic tasks, as well as lab management skills (i.e., personnel, grant-writing, communication). Research Plan: Spatial navigation is central to adaptive behavior, underlying our ability to trade-off the exploitation of our current location with the exploration of novel ones. Beautiful work has detailed a number of spatial codes (e.g., place and grid cells) in the hippocampal formation, yet we (1) lack a normative framework accounting for the complexities of natural navigation, (2) do not understand how spatial codes from the hippocampal formation interact with cortex, and (3) have focused on understanding how we build internal models of the world around us, while neglecting its starting point – ourselves. During the K99 phase of the award, I will develop a naturalistic navigation task in virtual reality where rodents will be required to disambiguate complex signals. These animals will be trained to integrate velocity signals derived from motion across their retina (i.e., optic flow) into a position estimate, in order to path integrate to the location of a latent target. Then, they will be tested in a novel situation, one where optic flow may be caused by self- and/or target-motion. I expect animals to behave in line with Bayesian Causal Inference (BCI) – a canonical computation wherein estimation biases emerge during small, but not large, signal disparities (i.e., when observers operate under the incorrect internal model). Further, I will broadly map neural activity throughout the rodent’s brain during BCI by leveraging novel large-scale neurophysiology techniques. During the R00 phase of the award, I will directly manipulate the subjective sense of self-location – the initial condition for navigation - and measure this phenomenology in rodents via the task developed during the K99. Beyond establishing BCI as a fundamental computation guiding naturalistic navigation, I expect the proposed project to inform next-generation therapeutics for disorders of inference, such as Autism and Schizophrenia.

NIH Research Projects · FY 2026 · 2023-05

PROJECT SUMMARY Background: Alcohol use disorder (AUD) has a lifetime prevalence of nearly 30%, with 14.4 million adults in the US currently in need of treatment. Even with treatment, 50-80% of individuals relapse within a year. Mechanisms underlying recovery are still not well understood, specifically individual differences underlying relapse risk. Preliminary data: The work of others and our preliminary data support the involvement of at least three neuro-behavioral mechanisms in the maintenance of AUD: 1) reward reactivity, 2) aversive reactivity and 3) executive control. Using a data-driven machine learning approach in a non-clinical community sample (N=1204; 46% male), we demonstrated that the top predictors of alcohol abuse constituted independent, additive factors of this three-domain model. Our preliminary analyses on subtyping in chronic poly-drug users (N=40; 75% male) and individuals with past AUD (N=74; 32% male), demonstrated that data-driven machine learning approaches can be used to study individual differences in these multi-factorial impairments. We found three distinct ‘subtypes’ in AUD: a “reward drinker” type (increased reward reactivity), a “relief drinker” type (increased aversive reactivity) and a “low functioning drinker” type (low executive control). Goals and Hypothesis: The immediate goal of this project in AUD is to develop a sparse personalized relapse prediction tool that can be employed in a treatment setting to continuously track relapse risk over time. The long-term goal is to determine if relapse prevention interventions can be personalized. The underlying hypothesis is that different combinations of independent factors underlie AUD maintenance and relapse per individual. We will test this by Aim I: determining individual differences in function on three domains (reward reactivity, aversive reactivity, executive control), and Aim II: evaluating the predictive power of subtype- or domain-specific relapse prediction models versus an AUD-general model, to determine their respective clinical utility. Specific Aims: In Aim 1, we will assess individual differences underlying AUD across the three domains of interest using a multi- method approach (personality, neurocognition, clinical assessments, task/resting fMRI brain function) in a large treatment cohort (N=200 AUD, 2-5 weeks into treatment, >40% female; N=100 controls). In Aim 2, we will follow our sample clinically (+6, +12 months) and employ machine learning methods to evaluate if the patterns of impairments underlying relapse risk are distinctly different between individuals. Innovation: This study provides a) a systematic, multi-method assessment of the individual heterogeneity in the neuro-behavioral mechanisms underlying AUD; b) an application of big data analytical approaches for relapse prediction to an AUD dataset; and c) the development of sparse yet highly informative personalized relapse prediction tools. Summary: This study will pave the way for the development of personalized relapse prediction tools that track relapse risk in AUD over time. This can ultimately lead to the development of personalized treatment approaches, with the potential to dramatically transform the current treatment landscape.

NIH Research Projects · FY 2026 · 2023-05

PROJECT SUMMARY/ABSTRACT Multi-organ failure as a result of the systemic inflammatory response to injury (SIRS) is the leading cause of late complications and death after severe trauma and burn injury. Despite decades of research, therapeutics that limit the SIRS response following injury remains an unmet clinical need. Emerging research points to the contribution of human-specific differences in our immune system that distinguish the human injury response from that observed in other species. These species-specific differences may explain why therapies that are widely successful in animal models used for preclinical research fail in human clinical studies. Uniquely human genes (UHGs), with expression that frequently tracks to human immune cells, may account for some of these differences in human SIRS after injury. The overarching goal of my research program is to systematically define factors that distinguish the human immune response from other species, characterize the contribution of UHGs to human SIRS, and understand how these genes effect therapeutics that target anti-inflammatory signaling pathways. To this end, we have recently discovered a novel role for the uniquely human CHRFAM7A gene that is a variant of the conserved α7 nicotinic acetylcholine receptor (α7nAchR) that mediates cholinergic anti-inflammatory signaling. In addition to decreasing the ability of therapeutics to target the α7nAchR, unexpectedly, we have demonstrated that human CHRFAM7A expression functions in transgenic mouse mice to cause increased monocyte mobilization to lung and decreased acute lung injury in a model of severe burn injury. Our research focus in this MIRA proposal is to identify the function and mechanism of action of UHGs that are highly expressed in monocytes/macrophages and relevant in the injury response. To demonstrate the functional relevance of UHGs, we will use a combination of precision animal models, genetic approaches in human induced pluripotent stem (iPS) cells, and clinical samples from trauma and burn patients. In this research program, we propose to 1) test cell-specific UHG function in an animal model of severe injury; 2) develop an iPS cell system for mechanistic studies in human macrophages; 3) determine how UHGs alter the effect of therapeutics that target anti-inflammatory signaling pathways; and 4) evaluate how relative UHG expression alters the inflammatory phenotype of monocytes from injured patients. Understanding how UHGs make the human immune response to injury unique may allow for the development of novel therapeutic interventions aimed at modulating SIRS and decreasing organ dysfunction after severe trauma and burn.

NIH Research Projects · FY 2026 · 2023-05

SUMMARY Dysregulation of inhibitory G protein-dependent signaling, including signaling controlled by the GABAB receptor (GABABR), is implicated in many neurological disorders and diseases. In the hippocampus (HPC), GABABR exerts much of its inhibitory influence by activating G protein-gated Inwardly Rectifying K+ (GIRK/Kir3) channels and by inhibiting Adenylyl Cyclase (AC). We have shown that GABABR-GIRK signaling in the HPC is modulated by Regulator of G protein Signaling (RGS) proteins of the R7 sub-family, which sharpen the timing and dampen the sensitivity of this signaling pathway. The R7 RGS sub-family includes RGS6 and RGS7, which form stable complexes with G5. Genetic ablation of RGS7/G5 complexes in mice profoundly alters GABABR-GIRK signaling, disrupting synaptic plasticity and HPC-dependent behaviors. R7 RGS/G5 complexes can also associate with adaptor proteins, including the small palmitoylated protein R7BP and an orphan Class C GPCR (GPR158). Our recent work suggests the intriguing prospect that discrete R7 RGS/G5 complexes, together with R7BP and GPR158, play an essential role in the selective routing of GABABR signals to GIRK channels or AC. These findings fuel our central hypothesis that R7 RGS/G5 complexes and their adaptors orchestrate the assembly of inhibitory "signalosomes” – distinct physical and/or functional arrays of receptors, G proteins, RGS and related proteins, and effectors – to ensure the dedicated and selective regulation of effector enzymes and ion channels by GABABR. To test this innovative hypothesis, we propose a multi-disciplinary program with two AIMs: 1) Identify mechanisms mediating GABABR-effector signaling dynamics and compartmentalization. We will employ loss- and gain-of-function/rescue genetic manipulations in HPC pyramidal neurons, along with electrophysiological and real-time optical assessments of GABABR-effector signaling, to test the prospect that discrete R7 RGS/G5 complexes and their adaptors, working in concert with their inhibitory G protein substrates, orchestrate the assembly of distinct and dedicated GABABR-GIRK and GABABR-AC signalosomes. 2) Interrogate the organization of the GABABR-GIRK signalosome. Leveraging insights derived from our recent crystal structure of RGS7/G5 and associated predictive modeling, we will use cell-based biochemical and optical reporter assays, as well as orthogonal biochemical reconstitution approaches, to test hypotheses related to interactions and interfaces among GABABR-GIRK signalosome elements. Structural insights into GABABR- GIRK signalosome assembly will then be exploited to probe the functional relevance of specific interactions. Summary: Our efforts will reveal whether R7 RGS/G5 complexes and adaptors orchestrate selective signaling between GABABR and its effectors. This project will yield new mechanistic insights into the functional and physical compartmentalization of GABABR-effector signaling, knowledge that can be used to design novel therapeutic interventions for neurological disorders and diseases characterized by aberrant GABABR-dependent signaling.

NIH Research Projects · FY 2026 · 2023-05

Project Summary/Abstract Children with developmental language disorder (DLD) demonstrate deficits in language compared to unaffected peers from similar language-learning environments. Conventional assessment methods for DLD capture a child’s current skill level in one language; in the U.S., such assessments are available only in English or Spanish. These methods fail sequential bilingual children, who speak a minority language at home and are subsequently exposed to English, because they confound language-learning ability with prior language- learning experience. Assessments that index language-learning ability are sorely needed, especially for sequential bilingual children whose home language is not Spanish. This project evaluates two novel approaches to language assessment for sequential bilingual children: dynamic assessment and processing-based assessment. Dynamic assessment methods offer language teaching during the assessment and evaluate the child’s response. Processing-based methods assess underlying skills that may drive language learning. This project rigorously examines the psychometric properties of dynamic and processing-based assessments within a diverse group of sequential bilingual children entering English-based schooling. We will recruit 165 children aged 4 years, 10 months to 6 years, 2 months who speak any non-English language at home. The first study timepoint will occur as children enter the school year and include an experimental assessment battery with dynamic assessments at the narrative and morpheme levels and both linguistic and non-linguistic processing- based assessments. The assessment battery will be repeated 1 month later. Aim 1 establishes psychometric properties of the experimental assessment battery, including internal consistency and test-retest reliability, factor structure, and concurrent validity. Aims 2 and 3 then use a longitudinal design to examine predictive validity. Children will be followed over two academic years, with re-assessment every 4 months (totaling 5 growth timepoints following the initial assessment and test-retest timepoint). At each growth timepoint, children will complete an English language sample and parents and teachers will be interviewed to assess the language environment and presence of concerns regarding the child’s language. We will construct models of English growth that account for child-external factors (e.g., language environment, SES) and then determine how well the experimental dynamic and processing-based assessment battery predicts this growth (Aim 2). We will then identify a group of children at high risk of DLD based on the combination of poor English growth and parent or teacher concern regarding language development. Aim 3 will examine the sensitivity and specificity of the initial assessment battery for predicting DLD. Project results will demonstrate whether multiple dynamic assessment and processing-based assessment tools can reliably and validly measure language-learning ability across variable language-learning environments. For millions of children who speak home languages other than English, this project represents a critical step in validating language assessment tools.

NIH Research Projects · FY 2024 · 2023-05

Human T-cell leukemia virus (HTLV-1) has been estimated to infect 15-20 million individuals worldwide and is known to be the etiological agent of an adult T-cell leukemia/lymphoma (ATLL), an inflammatory disease syndrome known as HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP), and pathologies of the lung, skin, eyes, and thyroid gland. HTLV-1 is notorious for being extremely difficult to propagate in cell culture, which has prohibited rigorous analyses of virus replication, including the steps involved in retrovirus assembly. HTLV-1 spread is known to be heavily reliant on virus infection involving cell-to-cell contacts that form what is termed the virological synapse (VS), which represents the primary means for virus spread, including events associated with oral transmission. While HTLV-1 has been previously studied in regard to virus spread via cell-cell contacts, a significant knowledge gap exists regarding the nature of virus particle assembly and transmission via the VS. In general, virus particle spread through cell-cell contacts increases the likelihood of an infection event of virus particles that may possess low particle infectivity in cells without formation of VS. Previous studies have indicated that a low proportion of mature HTLV-1 particles possess an intact capsid core, suggesting that aberrant particle morphology could help to explain the poorly infectious nature of cell-free HTLV- 1. In order to address the current knowledge gap in the field, we propose in this exploratory application to develop workflows for state-of-the-art bioengineering and quantitative imaging technologies that hold high promise in being applied to the efficient study of HTLV-1 particle assembly and spread at the VS. First, we will develop a workflow for the use of cell micropatterning technology in order to reproducibly and efficiently create cell-cell contacts and investigate the role virus budding in virus spread. Second, we propose to establish an efficient workflow in which we can view cell-cell contacts utilizing high-resolution cryo-correlative light and electron microscopy in order to investigate the role of host cell proteins in virus assembly at cell-cell contacts. Development of these workflows will allow for quantitative analysis of virus particle biogenesis at cell-cell contacts. These technologies have broad applicability in virology and the success of this research will be applicable to a variety of questions regarding virus replication and virus-host cell interactions.

NIH Research Projects · FY 2026 · 2023-05

Our goal is to develop small-molecule drugs for treatment of skeletal muscle disorders related to dysregulation of intracellular calcium, focusing on specific proteins in the sarcoplasmic reticulum (SR). Each Aim starts with the design of fluorescent biosensors (specific SR proteins labeled with fluorescent donor and acceptor), to be used in high-throughput screening (HTS) of small molecules. A key innovation is our recently developed HTS approach based on fluorescence lifetime (FLT) detection of protein structural changes by fluorescence resonance energy transfer (FRET). Our combination of FRET biosensor engineering with unique FLT detection has produced an unprecedented combination of sensitivity, specificity, speed, and precision in protein structure-based studies of mechanism for drug discovery. We previously validated this approach through applications to cardiac muscle. We now focus on skeletal muscle, targeting the two key SR proteins involved in Ca regulation, the Ca release channel (RyR1) and the calcium pump (SERCA1a). Aim 1: Targeting RyR1 leak reduction. Our biosensor is based on FRET between two regulatory proteins (FKBP12.0 and CaM) bound to RyR1. In pilot screens, we have shown that this FLT-based FRET assay can detect small molecules that restore aberrant RyR1 function, in which the Ca channel leaks Ca from the SR into the cytoplasm, inducing myopathies. We will carry out larger-scale screening, to identify new drug candidates, then use cellular and in vivo muscle assays to test the reversal of undesirable calcium leak in fibers and mice. Aim2: Targeting SERCA1a activation. We seek a complementary solution to combat Ca leak – enhancing SERCA1a activity to pump Ca back into the SR lumen. This approach also targets factors (e.g., mutation or oxidation) that impair SERCA1a activity. We will use two complementary approaches, building on our previous studies with SERCA2a (cardiac), with fluorescent biosensors expressed in live cells. (A) We will use an intramolecular FRET biosensor (donor and acceptor attached to different domains of SERCA1a), to screen a small-molecule library to detect compounds that bind to SERCA, alter enzyme structure, and activate Ca transport. (B) We will use an intermolecular biosensor, with donor on SERCA1a and acceptor on the SERCA1a regulator sarcolipin (SLN), to detect compounds that activate the enzyme by uncoupling the inhibitory effects of SLN. We will evaluate potency and efficacy of drug candidates, using assays on myofibers and muscles, both in vitro and in vivo in mouse models including pre-clinical longitudinal drug testing. We have assembled a multi-PI team with complementary expertise and decades of successful collaboration, led by David Thomas (SERCA1a, FLT-FRET), Razvan Cornea (RyR1, biosensor engineering), and Dawn Lowe (skeletal muscle functional analysis). We will also be joined by collaborators with complementary expertise in medicinal chemistry (Aldrich) and myofiber Ca assays (Launikonis), and two consultants with unique expertise on animal models of disorders in muscle Ca regulation (Dirksen and Hamilton).

NIH Research Projects · FY 2026 · 2023-04

Abstract Cryptococcal meningitis is the most common adult neuroinfection in sub-Saharan Africa and causes ~15% of AIDS-related mortality globally. In the weeks prior to onset of meningitis, cryptococcal antigen (CrAg) is detectable in the blood, and is a predictor of meningitis and death. CrAg screening in plasma is an effective public health strategy to identify persons with CD4<200 cells/mcL at high risk of meningitis and death. In a randomized trial of 2000 persons with HIV, CrAg screening and preemptive fluconazole treatment yielded a 28% survival benefit over standard-of-care. As a result, the World Health Organization and U.S. guidelines recommend CrAg screening. Yet despite the survival benefit seen with CrAg screening and preemptive therapy, 25% of initially asymptomatic CrAg+ persons treated with fluconazole still die, even with HIV therapy. From 4 prospective cohort studies of CrAg+ persons in Africa, we have determined that as plasma CrAg titer increases, mortality increases, despite fluconazole therapy. Among asymptomatic CrAg+ persons, disseminated cryptococcosis is the most commonly identified cause of death. We posit that subclinical disseminated early neuroinfection in the brain parenchyma accompanies high CrAg titers, and fluconazole is inadequate therapy. More effective treatment is critically needed to reduce mortality in CrAg+ persons. For symptomatic cryptococcal meningitis, amphotericin B is the most effective antifungal; fluconazole alone is inadequate. Recent randomized trial data reported single dose of liposomal amphotericin (AmBisome) 10 mg /kg with flucytosine (5FC) and fluconazole is as effective and less toxic than the traditional 7-day amphotericin + 5FC for meningitis. We hypothesize that AmBisome (10mg/kg x1), when combined with fluconazole, will be more effective than fluconazole monotherapy for asymptomatic CrAg+ persons. We have enrolled 244 CrAg+ persons in the initial phase II of a phase II/III randomized trial to demonstrate safety and feasibility of this enhanced regimen in Uganda, and now we seek to complete the phase III trial in order to test efficacy. The objective of this application is to assess the efficacy of AmBisome plus fluconazole to prevent cryptococcal meningitis and death. We will complete a randomized clinical trial of 600 CrAg+ persons (i.e. 356 more participants) to determine if preemptive therapy with AmBisome (10mg/kg x1) plus fluconazole for CrAg+ persons will improve cryptococcal meningitis-free 6-month survival compared with the current standard of fluconazole monotherapy (Aim 1). In Aim 2, we will determine if neurocognitive outcomes in CrAg+ persons preemptively treated with AmBisome with fluconazole are superior to outcomes with fluconazole monotherapy. Finally, in Aim 3 we will evaluate the cost and cost-effectiveness of AmBisome with fluconazole preemptive treatment in CrAg+ persons. Findings from this trial will impact U.S. and international HIV guidelines on the optimal prevention of cryptococcal meningitis, in order to reduce mortality in persons living with HIV.

NIH Research Projects · FY 2026 · 2023-04

While short-term outcomes following organ transplant have greatly improved with the development of more effective immunosuppression, long-term outcomes remain problematic with a significant number of patients developing diabetes, accelerated heart disease as well as increased rates of cancers and infections. For decades transplant researchers and clinicians have sought to develop strategies to induce immune tolerance to transplanted organs and avoid the requirement for life-long immunosuppression. It is likely that any successful tolerance regimen will incorporate targeted immunosuppression strategies like costimulation blockade. One of the most important interactions is the CD40-CD154 pathway. We will evaluate novel, clinically relevant therapeutics targeting either CD40 or CD154 and explore their role in facilitating tolerance. Based on our recent publication showing that CD11b is a novel alternate receptor for CD154 during alloimmunity, we will also test novel nanotechnology to block the CD154:CD11b interaction, an important mechanism of cross-talk between the innate and adaptive immune responses during transplantation. Moreover, based on our published data showing that memory T cells are a potent barrier to transplantation tolerance, we will determine the role of OX40-OX40L blockade to control alloreactive memory CD8+ T cells and promote a pro-regulatory environment. In addition, we will assess the importance of the VISTA pathway on the induction of donor-specific tolerance using an agonistic anti-human VISTA antibody. VISTA has a dual role in negatively regulating antigen-specific T cell responses while also impacting the innate immune response by inhibiting ischemia reperfusion injury, monocyte activation and neutrophil migration thereby suppressing the early inflammatory response. We will also investigate the contribution of IL-1 and the inflammasome on tolerance induction and employ clinically applicable therapeutics to control early inflammation during the induction of tolerance. The combination of cellular therapy and costimulation blockade is a powerful strategy to promote donor-specific tolerance. Myeloid derived suppressor cells (MDSCs) have inherent immunosuppressive properties and have been used to facilitate tolerance. They modulate innate immunity and inhibit T cell activation and effector cell function while also promoting regulatory T cell expansion for maintenance of long-term donor specific tolerance. Apoptotic donor leukocytes (ADLs) represent another promising cellular therapy that has been proven to control alloreactive T cells and promote donor specific T cell deletion. We will use donor bone marrow derived MDSCs or ADLs in combination with the above novel therapeutics to promote tolerance in nonhuman primate kidney transplantation.

NIH Research Projects · FY 2026 · 2023-04

Systemic inflammation and oxidative stress are common in patients with AF. In atrial cardiomyocytes (CMs), AF can be precipitated by NLRP3 inflammasome activation and IL-1β secretion. Since we have established cardiac NLRP3 activation and IL-1β can lead to AF, we will study upstream modulators of the cardiac NLRP3 inflammasome that can be manipulated to reduce AF risk in DM. We have found that enzymes producing pro-inflammatory molecules are elevated and inflammation resolving molecules are reduced in atria from humans and mice with DM. Specifically, we have found increased 12- lipoxygenase (12-LOX, encoded by ALOX12), an enzyme that processes arachidonic acid (AA) to pro- inflammatory metabolites in humans and mice. In humans and mice, we have found that cardiac pro-resolving lipid mediators (SPMs) are reduced, leucine-rich repeat containing G protein-coupled receptor 6 (LGR6, encoded by LGR6), a recently described receptor of SPMs, is downregulated, and 15-hydroxyprostaglandin dehydrogenase (15-PGDH; encoded by HPGD), an enzyme in the inactivation of SPMs, is increased in DM atria. Hypotheses to be tested: Since SPMs can reduce NLRP3 activation, this application explores whether DM- associated AF risk can be mitigated by enhancing SPM signaling by reducing inflammatory lipid mediator production (12-LOX inhibition), enhancing SPM signaling (upregulation of LGR6), or reducing SPM degradation (downregulating of 15-PGDH). Specific aims: Aim 1: Determine whether inhibition of cardiac 12-LOX upregulation can reduce atrial NLRP3 activation and AF burden in DM. Specific Aim 2: Determine whether upregulation of cardiac LGR6 can reduce atrial NLRP3 activation and AF burden in DM. Specific Aim 3: Determine whether downregulation of cardiac 15-PGDH can reduce atrial NLRP3 activation and AF burden in DM. Significance: This application explores new treatment paradigms of encouraging inflammation resolution to prevent DM-induced AF. Using parallel experiments in humans and mice will provide mechanistic insights and strengthen clinical relevance. A focus on prevention rather than treatment is novel and could prevent significant morbidity associated with AF onset.

NIH Research Projects · FY 2026 · 2023-04

PROJECT SUMMARY/ABSTRACT A watershed moment in decades of research in Alzheimer's disease (AD) is the discovery that targeting certain molecules in microglial cells can improve a hitherto unknown functional mechanism in these cells, and arrest cognitive decline. The first in class of microglial molecular targets for AD therapy is TREM2. TREM2 expression is upregulated in microglia in the AD brain, the microglia changes from a homeostatic state to something known as damage-associated microglia (DAMs) as defined by transcriptomics, and the loss of TREM2 prevents DAM transition while accelerating disease progression. Notably, the loss of TREM2 prevents the upregulation of another microglial molecule - AXL. Whether AXL has a critical effector function in the DAM-mediated thwarting of cognitive decline remained heretofore unknown. We have discovered that augmenting AXL leads to the arrest of cognitive decline in a mouse model of AD. There is a potential third player in this axis - MERTK. MERTK has been implicated by an independent study (Huang et al., Nature Immunology, 2021) in the phagocytic engulfment of filamentous A[l by microglia and its eventual compaction into harmless dense core plaques. Here we propose a systematic approach to understand this still nebulous process of microglia-mediated arrest of cognitive decline. First, we will use mouse genetics to investigate the requirement of sequential involvement of TREM2, followed by AXL and likely subsequently by MERTK in microglia-mediated arrest of cognitive decline in mouse models of AD through behavioral tests and electrophysiological assessment of learning and memory. Second, we will correlate these genetic epistasis-associated functional changes in learning and memory to corresponding transcriptional state of microglia as assessed by single nucleus RNA sequencing. Our third aim is to evaluate cellular, subcellular, morphological and neuronal network level brain functional changes, including AD neuropathological hallmarks such as amyloid plaques and tau phosphorylation, as well as microglial functions such as phagocytosis and/or plaque barrier formation. We hypothesize that a TREM2, AXL and MERTK triad functions sequentially to engineer a beneficial microglia state, which in turn counters AD-associated ill-effects that manifest as cognitive decline. Therefore, augmenting the function of this triad would restrain cognitive decline and preserve brain health in AD. Our study could lead to the development of multivalent engagement of microglial molecules TREM2, AXL and MERTK, for novel therapeutics in AD.

NIH Research Projects · FY 2026 · 2023-04

Abstract Ewing sarcoma survival has not improved in decades despite long knowledge of its singular driving somatic mutation, the pernicious EWS-FLI1 fusion oncoprotein. As an aberrant transcription factor EWS-FLI1 alters expression of thousands of genes enacting a complex program toxic to most cell types, so much so that the cell of origin for Ewing sarcoma is still a matter of debate. Ewing sarcoma occurs at starkly higher rates (roughly 10- fold) in children of European ancestry compared to those with primarily African ancestry, while Latino children have roughly ⅔ and Asian children ½ the risk of Ewing sarcoma than do white children. Ancestry is in fact the strongest known risk factor for Ewing sarcoma, but the molecular basis of these differences in risk have been investigated only sparingly. As the mechanisms by which EWS-FLI1 interacts with the genome have become clear, it is feasible with an appropriate model to investigate how genomic ancestry in general and at specific loci modulates EWS-FLI1 activity including its downstream effects on epigenetic and transcriptional programs. To this end we have devised a strategy to introduce EWS-FLI1 in derivatives of induced pluripotent stem cells (iPSC) in order to characterize downstream molecular and functional consequences of EWS-FLI1 expression. Using accessible variant array data from several large stem cell repositories we identified banked iPSC lines derived from individuals with a range of European, African, and Amerind ancestry. We will introduce EWS-FLI1 expression at intermediate stages of development relevant to Ewing sarcomagenesis—namely neural crest cells and mesenchymal stem cells. Measures of functional tolerance and molecular state will be compared to corresponding samples from subjects with solely European ancestry. Globally we will examine whether gene expression and chromatin state exhibits similarity to the Ewing expression signature in proportion to European ancestry percentage. Genome-wide chromatin occupancy of EWS-FLI1 will be profiled and its relationship to local ancestry defined using long-read sequencing. Genes that are differentially influenced by EWS-FLI1 in “permissive” (European ancestry), “moderately permissive” (Amerind ancestry) and “impermissive” (African ancestry) genomes will be considered targets of potential therapeutic value and will undergo validation using CRISPR/Cas9 genome engineering and functional assays. The resulting data will represent the first and only effort, to our knowledge, to take advantage of the known differences in risk for Ewing sarcoma by ancestry to study EWS-FLI1 binding and downstream effects. In addition, we will make available the genome-scale data produced by our study and freely distribute our iPSC models for wider use by Ewing sarcoma researchers.

NIH Research Projects · FY 2025 · 2023-04

PROJECT SUMMARY Chronic traumatic encephalopathy (CTE) is a neurodegenerative disease associated with repeated mild traumatic brain injury (TBI). CTE is among the many neurodegenerative diseases characterized as tauopathies, wherein the protein tau, which is usually associated with microtubules in the axons of neurons, becomes separated from microtubules, initiating a degenerative cascade and leading to eventual neurofunctional loss. There are currently no pharmacological treatments available for CTE patients, so any treatment that could limit or reverse tau-associated dysfunction would have an important impact on TBI patient outcomes. Moreover, given the similarity between CTE and other tauopathies, insights into CTE treatment could be broadly applicable to other common neurodegenerative diseases. We have recently developed an in vitro model that directly links mechanical injury to tau pathology in cultured neurons. One notable outcome from our prior studies is that we found that synaptic dysfunction (measured using patch clamp) is correlated with mislocalization of tau to dendritic spines (measured using fluorescent imaging). This result suggests that this relatively simple image-based readout could be used for screening the effects of pharmacological agents on the functional progression of trauma-induced tauopathy. There are currently no high-throughput in vitro models for screening the effects of pharmaceuticals on trauma-induced tauopathy. We have developed several cell stretching systems, both for neurotrauma and other applications. However, all of these systems are far too low-throughput for performing drug discovery studies. Thus, our goal is to scale up our current in vitro neurotrauma model to a high-enough throughput for drug screening studies. We will design a new stretchable multi-well plate for neuronal cell culture and a new high strain rate stretcher that can apply trauma-like loads to the cells in the plate. In addition, we will employ machine learning based algorithms to quickly and efficiently analyze the data collected from our new device. Finally, we will use the device to test inhibitors known to be effective against other aspects of neuronal injury affect tau mislocalization